The present invention is directed to the creation of two novel cell lines, the GABAergic hNT2.17 and serotonergic hNT2.19, synthesizing and releasing the neurotransmitters GABA and serotonin (5HT), both derived from the human NT2 cell line, and which are useful for the treatment of pain and spasticity and other traumatic or neurodegenerative injuries to the nervous system. These cells are stably transplanted into host rodent animals in models of pain and spasticity.
Chronic pain and spasticity constitute some of the significant problems following spinal cord injuries (SCI), interfering with rehabilitation and daily activities, which in turn leads to significant complications in these patients. However, there are no effective, completely safe therapies for the reduction of these conditions (and other related to neural trauma and degeneration).
Oral baclofen, the GABA-B receptor agonist, has often been the drug of choice for spasticity (Reference 1) due to spinal cord trauma, and baclofen and opioids have been used for the relief of chronic pain (Reference 2). However, oral baclofen is often ineffective at non-toxic doses, because large doses are required to cross the blood-brain barrier and can lead to subsequent CNS effects (Reference 3), usually not well tolerated by spastic patients. Intrathecally administered baclofen is currently FDA-approved for those patients with spinal cord injury who are refractory or can not tolerate oral baclofen, but spinally-delivered baclofen via programmable implantable electronic pumps is also fraught with problems, such as high costs, catheter twisting, infection at the implant site, overdosing and the development of tolerance with increasing doses of baclofen required for relief (References 4 and 5). Furthermore, the efficacy of baclofen pumps is mainly in terms of lower extremity spasticity and the treatment has little effect on upper extremity spasticity. Intrathecal morphine has been used via electronic pump and has been prescribed for use in neuropathic pain, but often has complications of tolerance, overdose, and problems related to the pump itself (References 6-8).
Transplants of primary cultured cells near the dorsal horn of the spinal cord that release peptides and neurotransmitters have offered a new direction in the treatment of chronic pain (Reference 9). However, primary cells are difficult to obtain, non-homogeneous, and would require that each batch be tested before clinical use. Transplantation of immortalized cell lines genetically-modified to release neuroactive antinociceptive peptides (Reference 10), inhibitory neurotransmitters (References 11 and 12) and neurotrophins (Reference 13) in chronic pain, and to upregulate inhibitory neurotransmitter synthesis (Reference 14) offers a renewable source of cells that can act as cellular minipumps, are able to respond to the microenvironment of the cord, and should reduce or eliminate side effects associated with the large doses of pharmacologic agents required for centrally-acting pain-reducing agents. A naturally-immortalized human embryonal carcinoma cell line, NTera2cl.D/I (NT2), differentiates irreversibly into several morphologically and phenotypically distinct cell types, which include terminally differentiated postmitotoc CNS neurons (Reference 15). Successive re-plating of retinoic acid-treated NT2 cells, in the presence of growth inhibitors, results in the isolation of purified human neurons (Reference 16), that have been extensively characterized and tested in vivo in a number of animal models of traumatic injury and neurodegenerative disease (Reference 17). The potential application of NT2 neurons in cell transplantation therapy for CNS disorders, and their use as vehicles for delivering exogenous proteins into the human brain for gene therapy, has been recently demonstrated (Reference 18). Such NT2 neurons are being currently used in Phase-II clinical trials for the treatment of stroke, and have been approved by FDA for such trials (References 19 and 20). However, such NT2 cells contain a variety of neural phenotypes, and would provide a plethora of neuroactive substances if used for the specific treatment of problems such as pain or SCI-associated spasticity.
U.S. Pat. Nos. 5,082,670, 5,175,103, 5,792,900, 6,162,428, 6,214,334, and 6,254,865, are directed to the use of the parental human NT2 cell line for a variety of disorders, but none describe the creation or use of individual cell lines derived from the NT2 or NT2/D cell lines.
In order to ameliorate various neurological diseases, conditions, or disorders, the present invention creates two novel human neural cell lines, subcloned and derived from the human NT2 cell line, where each subcloned cell line synthesizes and secretes bioactive agents, such GABA or serotonin (5HT). Transplants of these secreting human neural cell lines are believed to reduce consequences of spinal trauma after transplant in or near the spinal cord.